Multiple drug resistant tuberculosis (MDRTB) has recently emerged as a serious public health concern. To develop better drugs to treat MDRTB and design improved diagnostic protocols, it is imperative that drug targets and drug resistance mechanisms of M. tuberculosis are defined. Our laboratory has investigated the mechanism of resistance to Isoniazid, rifampin and streptomycin in 53 clinical M. tuberculosis isolates. We have demonstrated that reduced sensitivity to INH often is associated with alterations in the catalase-peroxidase (kat G) locus or in the promoter of the inh A gene. Resistance to rifampin is conferred by mutations in the rpo B gene (RNA polymerase). Genetic alterations in the rpo L (ribosomal S12 protein) on the 16 sr RNA genes are primarily responsible for Streptomycin resistance in MDR tb strains. Furthermore, we established that multiple drug resistance in M. tuberculosis results from an accumulation of mutations in genes encoding drug targets and not from a novel pleiotropic resistance determinant.